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Question: answer questions 16 based on experimental pages from the first...

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Answer questions 1-6 based on experimental pages from the first two pages. Thanks!
29 LEAF DISC GROWTH ASSAY af growth involves a period of cell proliferation followed by a period of an increase in leaf area. The final surface area of a leaf is dictated both by Background: The process of cell expansion, leading to conditions including the availability of water, light and nutrients. The processes regulated by the leaf genetics and by environmental rate of leaf expansion is determined by biochemical and biophysical tissue, such as turgor pressure and osmotic gradients, a individual cell. In this lab, you will work in small groups and combine efforts from the entire class to explore that light, the solutes potassium and sucrose, and the inhibitors DCMU (a compo s well as the cell wall extensibility of an the roles nent of commercial herbicides) and abscisic acid (ABA) play in the process of leaf expansion Experimental Set-up The class will be divided into small groups to perform one of the two following experiments: Dark + 10 mM KCI and 1 mM Sucrose Light 10 mM KCI and 1 mM Sucrose Light+ 10 mM KCI, 1 mM Sucrose, and 106 HM DCMU Light+ 10 mM KCI, 1 mM Sucrose, and 10 HM DCMU Light+ 10 mM KCl, 1 mM Sucrose, and 104 uM DCMU Experiment 2 Dark +10 mM KCI and 1 mM Sucrose Light +10 mM KCI and 1 mM Sucrose Light +10 mM KCI and 1 mM Sucrose 10-6M ABA Light +10 mM KCI and 1 mM Sucrose+10- M ABA Light +10 mM KCl and 1 mM Sucrose+ 10 -M ABA eparation of Stock Solutions: 1) The instructor will assign each group a stock solution to prepare and share with other groups ) Determine the concentration of the stock solution that you will need to prepare, keeping the following considerations in mind: -Each light/solution treatment will be contained within one petri dish containing 40 mL of solution. (What is the total volume of stock solution you will need?) You will need to make enough of the stock solution to share with another group. (Double the amount you are making for your group alone, then multiply to make enough for other groups) - Your stock solution should have a higher concentration than that of the treatment solution. Molar weight the number of grams needed to make 1 L at a concentration of 1 M
3) Once you have determined the volume and concentration of stock solution that you will be king, calculate the number of grams you will need of the chemical, weigh it out on the maki balance and place it in a beaker. Add dH0 up to a few milliliters less than the total volume needed. Add a stir bar to the beaker and place beaker on stir plate to dissolve the solute. Once it is dissolved, pour the solution into a graduated cylinder and add dH2O to the final volume. Pour stock solution into a bottle with lid. Label bottle with contents including the chemical name, concentration, and date made Note: if you are making a DCMU stock solution, you will first need to dissolve the DCMU powder in ethanol before adding water. A few milliliters of EtOH are all you need. Please do this in the fume hood in the inner lab. Ask your instructor if you are unsure. Once all the stock solutions have been prepared, you will need to calculate the volume of e that will go into the petri petri dish will be 40 mL. How much stock sucrose solution will concentration of 10 mM? How much stock KCI solution will you need to make a final Kc concentration of 10 mM, etc. Add dH20 to attain the final total volume of 40 ml 4) ach dishes as your leaf growth solution. Again, the total volume in each you need to make a final sucrose A very useful equation: 5) After the petri dishes have been labeled with solution, light treatment and group name, and filled with the appropriate growth solution, leaf discs can be made. Place a few paper towes on the bench and moisten with dH20. Select several still expanding leaves from the bean plants provided. Your instructor will show you an example of an expanding leaf. Place the leaves on the paper towel and using a cork borer, make multiple leaf discs and place them in the petri dishes using a pair of forceps. You should make 10 leaf disks per treatment. Be sure to note the size of cork borer you use as this will be the standard size against which any changes due to growth will be measured. 6) Take petri dishes to the light treatments as indicated by your instructor. the following day, you will measure the new area of the leaf discs and compare the change in size to the standard. Average the values for each treatment and share with the class. 7) After 24 hours, they will be moved to a refrigerator to arrest growth. When you return to class
31 Leaf disc growth assay Pre-Lab Questions 1. Calculate how much sucrose you will need to weight out to make one liter of a stock solution of 100 mM sucrose. (2 points) a o.IM34.23 A2. What concentration of ABA would make a useful stock solution for the class to use? (2 points) 3. What is the role of potassiun in these experiments? (1 point 4. What is the role of sucrose in these experiments? (1 point) 5. How will DCMU inhibit growth of the leaf discs? Be explicit. (2 points) reduction f NADP NADP and prevent tne for manon of oxygen fom prevents te protn qodiens fromeh nPrveie ians from onve 6. What light intensity should you use for the Light treatments? Why? (2 points)
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